Manufacturers and respective regulatory agencies strive to continually improve the safety and the quality of foods, pharmaceuticals, neutraceuticals, water and the environment. Under recent United States Department of Agriculture, Food Safety and Inspection Service (FSIS) directives and guidelines, many food producers in the United States have adopted sampling plans that involve testing, for example, of raw beef trims and/or ground beef for E. coli O157:H7, and ready-to-eat products for Listeria monocytogenes and/or Salmonella. Similar trends are observed in other countries, for products detained for domestic consumption and/or export. For example, the pharmaceutical industry spends considerable resources on sterility testing and environmental monitoring, and the water and wastewater industry and their respective regulatory agencies allocate considerable resources to monitor the quality of water (e.g., drinking, recreational and fisheries resources), wastewater (treated and untreated) and sludge (biosolids). Concern for the quality of air has resulted in monitoring of indoor air for the presence of microbial contaminants and pathogens. These sampling plans associated with such testing determine, for example, the disposition of products and the production lots, or result in decisions regarding the safety of water, receiving waters and recreational waters, and the impact of wastewater discharge, and quality of air.
The operative art with respect to such product “hold and release” monitoring efforts comprises various different sampling plans, typically pursuant to International Commission on Microbial Specifications for Food (ICMSF) guidelines for testing food products, FDA and USDA or EPA guidelines, or intuitive sampling plans. The common feature of these sampling plans is that several samples representing production lots, or a number of environmental samples, are combined into a combined production lot prior to sampling. For example, a typical plan for microbial pathogen detection calls for taking more than one sample, for instance ten (10) to sixty (60) sample pieces from a given sample lot (combined production lot), to form a composite sample for testing. The number of pieces taken to form the composite sample depends, under ICMSF or other regulatory guidelines, on the outcome of infection/poisoning (e.g., the severity), the level of hazards, and potential increase in the hazard levels due to storage. The same holds true in detecting microbial constituents and/or spoilage organisms. With respect to E. coli O157 testing, for example, a typical plan calls for taking 60 samples (e.g., sample pieces) from a production lot to form a single composite sample. With respect to Listeria monocytogenes, thirty (30) samples are typically taken from a production lot to form a composite sample. With respect to sterility testing, a typical plan may call for compositing 10-100 units of the product, and in the case of environmental sampling, 2-10 samples are often composited. Generally, the number of samples that are composited reflects: the overall sensitivity of the test method and the limit of detection that applies to the test unit; the ability to concentrate/enrich the composite sample to achieve the desired sensitivity; and the cost of sampling compared with the cost of analysis.
The costs associated with testing food are substantial, and consequently manufacturer's often resort to increasing the size of the “production lot” to be tested. For instance, the standard size production lot for trim testing for E. coli O157 in the U.S. beef industry is five (5) ‘combos’ (each combo weighing approximately 2,000 pounds), and for Listeria and Salmonella in ready-to-eat (RTE) products, it is one composite sample per production line per shift. A primary problem associated with the use of large size test lots is that a large quantity of products must be downgraded or destroyed when there is a positive finding of a pathogen. Additionally, because of the nature of combination ‘lotting’ (combining, for purposes of forming the test lot, several independent sub-lots that collectively span a long period of production time), it is very difficult to investigate the cause of the failure and pinpoint the source/cause, and remedial measures cannot be effectively taken. Furthermore, compositing a large number of samples may result in reduced sensitivity for the test unit and adversely effect the limit of detection.
Typically, once a composite sample is constructed (following the art with respect to particular testing) it is then enriched for the presence of a given pathogen/microbe by addition of appropriate amounts of enrichment media and incubation at an appropriate temperature for a given amount of time. If the pathogen/microbe is present, then it grows and multiplies under the favorable conditions of enrichment, thereby providing more material which can be detected by subsequent analysis. The composite sample is then tested by one or more available, art recognized methods including, for example, enrichment followed by immunoassay-based tests or PCR-based methods, or culturing of the organism of interest, or other DNA- or immunochemical-based methods.
The above-described prior art has several substantial deficiencies. First, even though the prior art production lot encompasses several production sublots (for instance 5 combo/pallets of products, or an entire shift of production), if a positive finding is obtained for the production lot (composite test sample), the entire production lot is rejected (all 5 combos/pallets, or one shift of production) and diverted to economically undesirable end uses (e.g., cooking or disposal), even though the pathogen of concern may be confined to only a limited portion of the production sublots (corresponding to one or two combos/pallets or an hour of production) comprising the production lot.
Second, the current art does not allow for retesting of any production sublots, primarily because microbial contaminants are unevenly spread throughout the products, and in many instances the levels of contamination are minute and may be present on very small portions of the products in each sublot. Therefore, once a composite sample is constructed if it tests positive all of the sublots are destroyed. The regulatory agencies do not allow for re-testing of the sublots, premised on the argument that after a production lot tests positive, negative test results obtained for production sublots, or even a new production lot composite sample, are meaningless since the microbial contaminants are not uniformly present throughout the products.
Third, the current protocols for collecting a composite sample representing a production lot often group together production sublots that are unrelated by virtue of product type or hours of production. This makes investigating the nature of the failure a difficult task even when relevant information is available. As an example, information is frequently available on a ‘per combo’ basis including (but not limited to) hour of production, vendor source of raw materials, production employees present, and operational status of microbial intervention process steps. However, when five unrelated combos/pallets are included in a production lot it is much harder to rationally analyze the available information since there is no way to determine which combo(s) contained the pathogen (as stated above, prior art re-sampling is not a viable option since it may not yield the same result). Likewise, for sterility testing, when the composite sample tests positive, the entire production will be disposed.
Therefore, there is a pronounced need in the art to implement novel and effective testing protocols that provide a greater measure of assurance that the food product is safe, that provide economic relief to the producer, and that allow for effective tracking of the contaminated lot(s) for remedial purposes (e.g., pinpointing the time of the contamination and determine the segment of production which was impacted).